Isolation and Characterization of Salt and Temperature Tolerant Plant Growth Promoting Rhizobacteria for Enhancing Tomato Plant Growth in Saline Environments

doi:10.21203/rs.3.rs-3402915/v1

The present study was focused on isolation, selection and characterization of salt and temperature tolerant plant growth promoting rhizobacteria from rhizosphere soil of tomato. A total of 46 isolates were developed on Ashby's Mannitol agar media out of 138 rhizobacteria. The medium was supplied with various NaCl concentrations, MgCl₂ concentrations, and pH levels to examine the tolerance to salt and temperature. All the forty six isolates showed promising salt tolerance. Out of forty six isolates, nine isolates were extremely salt tolerant (more than 5.4% salt tolerance limit). Two isolate were tolerant to NaCl concentration (2.10–3.6% salt). Nine isolates were moderately tolerant to NaCl concentration (0.09–1.50% salt). Twenty six isolates were moderately sensitive to NaCl concentration (0.15–0.60% salt). On the basis of the salt tolerance limit, rhizobacteria that promote plant growth in salt and temperature-tolerant environments were categorized. Out of forty six isolates, eight isolates showed growth at MgCl₂ concentration of 4.0%, two isolates showed growth at 3.0% MgCl₂ concentration, thirteen isolates showed growth at 2.0% MgCl₂, fifteen isolates showed growth at 1.0% MgCl₂ and eight isolates showed growth on 0.5% MgCl₂ concentration. Out of the 46 isolates, eight showed growth at pH of 9.0, whereas the other 38 showed development at pH of 7.0. The plant growth promoting activity test was positive for four isolates (Pseudomonas furukawaii STT-A8, Achromobacter sp. STTA-12, Agrobacterium pusense STT-A24, and Agrobacterium pusense STT-A39) out of the forty six total isolates. Pseudomonas furukawaii STT-A8, Achromobacter sp. STT-A12 Shown temperature tolerance limit at 60⁰c and Agrobacterium pusense STT-A24 and Agrobacterium pusense STT-A39 shown temperature tolerance limit at 50⁰c. The experiment was done in order to isolate, pick out, and characterize salt and temperature tolerant plant growth-promoting rhizobacteria that would be a crucial inoculum to enhance tomato plant growth and development in a salty environment. There is currently need to develop highly salt and temperature tolerant plant growth promoting rhizobacterial consortium.

Isolation

characterization

salt and temperature tolerant

plant growth promoting rhizobacteria

In dry and semiarid areas, salinity poses a severe hazard to agriculture (Rao and Sharma, 1995). Salinity has an impact on around 40% of all land on the planet. According to Egamberdiyeva et al. (2007), soil salinity is a possible issue in irrigated soils that can lower soil and water quality, restrict crop development, and result in the abandonment of agricultural property. Large active groups of bacteria known as plant growth promoting rhizobacteria (PGPR) are supported by the rhizosphere, which is the term for the thin layer of soil around plant roots and the soil that the roots occupy. According to Rangarajan et al. (2003), rhizobacteria that promote plant growth are known to quickly colonise the rhizosphere and reduce soil-borne diseases at the root surface. By promoting growth, these organisms can also be advantageous to the plant (Bloemberg and Lugtenberg, 2001). The most promising class of rhizobacteria that promote plant growth and are used in the biocontrol of plant diseases is Fluorescent Pseudomonas. They generate secondary metabolites such phytohormones, siderophores, volatile hydrogen cyanide, and antibiotics. These bacteria's capacity to stimulate plant development is mostly attributed to the production of antibiotics, siderophores, and indole-3-acetic acid.The Enterobacteriaceae family includes some of the genera of PGPR, including Azotobacter, Azospirillum, Pseudomonas, Acetobacter, Burkholderia, Bacillus, and Paenibacillus. The direct application of microorganisms to enhance plant development and prevent plant pests is still a fast developing research field. One of the initial stages in the pathogenesis of soil-borne bacteria is rhizosphere colonisation. The utilisation of microbial inoculants as biofertilizers, biocontrol agents, phytostimulators, and bioremediators depends on it as well. Pseudomonas spp. are frequently presented as examples of bacteria that colonise roots (Lugtenberg et al., 2001).The ability of these rhizobacteria to produce a variety of substances, such as phytohormones, organic acids, siderophores, atmospheric nitrogen fixation, phosphate solubilization, antibiotics, and some other unidentified mechanisms, has been variously attributed to their beneficial effects (Glick and Ibid, 1995). Commercial PGPR applications are being tried with good results. The success rate of field applications will be significantly improved, however, by a better knowledge of the microbial interactions that lead to plant growth improvements (Farzana et al., 2009). It is known that PGPR, root-colonizing bacteria can affect plant growth through a variety of direct or indirect processes. With PGPR, several chemical alterations in soil are connected. Numerous methods have been suggested by research to explain how plant growth-promoting bacteria (PGPB) affect nutrient intake, yield, and growth. Some bacterial strains mimic the manufacturing of plant hormones in order to control plant physiology directly, while others boost the availability of minerals and nitrogen in the soil to promote growth. In comparison to the uninoculated positive controls, the 14 halotolerant bacterial strains increased root length by between 5.2% and 47.8% and dry weight by between 16.2% and 43% in canola plants under salt stress (150 mM NaCl). Particularly, when compared to uninoculated salt-stressed canola seedlings, three of the bacteria—Brevibacterium epidermidis RS15, Micrococcus yunnanensis RS222, and Bacillus aryabhattai RS341—showed more than 40% increases in root elongation and dry weight. These findings suggest that specific halotolerant bacteria isolated from coastal soils may be able to improve plant growth in salt water environments by reducing ethylene production through ACC deaminase activity (Siddikee et al., 2010). As compared to a typical free diazotrophic PGPR, Azotobacter chroococcum EMCCN1458, tomato inoculation with the acquired Achromobacter sp. EMCC1936 considerably boosted vegetative growth, yield characteristics, and endogenous phytohormone concentration. Agrobacterium pusense significantly produces indole-3- acetic acid (86.95 mg/l), gibberellic acid (172.98 mg/l), and ammonia (42.66 mol/l) as a qualifying plant growth-promoting bacteria. Additionally, it generates siderophores, 1- aminocyclopropane-1-carboxylic acid deaminase, fixes nitrogen, makes biofilms, and effectively solubilizes zinc and inorganic phosphates from the soil( Kaur et al.,2022). The nitrogen fixation in plants may be impacted by a variety of environmental stressors. Salinity, water stress, soil pH, temperature, and heavy metals are some of its components (Kucuk and Kivanc 2008). According to Singleton et al. (1982), increasing the salt content could harm the rhizobacterial population. When tomato crops were infected with salt and temperature tolerant rhizobacteria, their capacity to grow and survive in saline environment was enhanced. Therefore, the isolation of salt and temperature tolerant rhizobacteria would be the most crucial inoculums to enhance plant growth and development in a saline environment.

Collection of rhizosphere soil samples of tomato

Total forty six (46) rhizosphere soil samples were collected from saline tract of four districts of Western Maharashtra, India viz., Ahmednagar, Nashik, Pune and Satara to isolate salt and temperature tolerant rhizobacteria. From each site, soil samples were randomly collected by dividing the field into different homogeneous units based on visual observation. At the sampling location, the surface trash was cleared away. The auger was driven to a plough depth of 15 cm, and the soil sample was drawn. Ten samples were collected from each field and placed in a polythene bag. The soil samples were thoroughly mixed, and roots, stones, pebbles, and debris were removed. The soil samples were air-dried by spreading them on a clean surface and allowed to dry naturally, avoiding direct sunlight and excessive heat. The bulk of the samples were reduced to one kilogram by thoroughly mixing the sample into four equal parts. Two opposite quarters were discarded, and the remaining two quarters were remixed. The method was carried out several times to get the required sample size. The final sample was collected in a clean cloth or polythene bag. The bag was labelled with relevant information such as the name of the farmer, location of the farm, survey number, previous crop grown, present crop, crop to be grown in the next season, date of collection, and the name of the sampler.

Serial dilution

Rhizosphere soil samples were immediately transported in sterile plastic bags to the department of plant pathology and agricultural microbiology laboratory and then were 1 g of each soil sample was dissolved in 9 mL of sterile deionized water before being serially diluted. To create a 10^− 1 dilution, a total of 100 µL of the stock solution was combined with 900 µL of sterile distilled water. This process was repeated until a 10^− 6 dilution was achieved. A 100 µL sample of each dilution was then plated onto Ashby's Mannitol agar. Before autoclaving, the initial media's pH was corrected to 7.00. Two days post incubation at 28 ± 2 ⁰C, the plates were observed and checked the growth of colony. Selected rhizobacteria isolates were purified, stored, and selected for further studies.

Assessment of salt and temperature tolerant rhizobacteria at different NaCl level.

To confirm the salt and temperature tolerance of rhizobacteria isolates, they were tested against different concentrations of NaCl salt and classified on the basis of salt tolerancy method given by Cardoso et al, (2014).For this, Ashby’s Mannitol agar media supplemented with 0.15, 0.3, 0.6, 0.9, 3.0 and 6.0 per cent NaCl. The plates of Ashby’s Mannitol agar media with above NaCl concentrations were prepared on which each rhizobacterial isolates were inoculated by streaking and incubated at 28⁰C for 2–7 days. After completion of incubation period, growth of salt and temperature tolerant rhizobacterial isolates were observed for growth and rated as +++: full growth, ++: scanty growth +: growth and -: no growth. The full growth rhizobacterial isolates were selected for further studies.

Assessment of salt and temperature tolerant Rhizobacteria on different MgCl ₂ concentration.

To confirm the salt and temperature tolerance of rhizobacterial isolates, they were tested against different concentrations of MgCl₂ salt. For this, Ashby’s Mannitol agar media supplemented with 0.5, 1.0, 2.0, 3.0, and 4.0% MgCl₂. The plates of media Ashby’s Mannitol agar with above MgCl₂ concentrations were prepared on which 46 isolates of salt and temperature tolerant rhizobacteria was inoculated on media by streaking and incubated at 28^oC for 2–7 days. After completion of incubation period, growth of salt and temperature tolerance rhizobacterial isolates were observed for growth and rated as +++: full growth, ++: scanty growth +: growth and - : no growth. The full growth rhizobacteria isolates were selected for further studies.

Assessment of salt and temperature tolerant rhizobacteria on different pH level.

To confirm the salt and temperature tolerant rhizobacterial isolates, they were tested against different pH levels in the medium. For this, Ashby’s Mannitol agar medium, pH was adjusted by using HCl and NaOH to 7.5, 8.00, 8.50 and 9.00. The plates of Ashby’s Mannitol agar with above pH levels were prepared on which each salt and temperature tolerant rhizobacterial isolates were inoculated by streaking and incubated at 28⁰C for 2–7 days. After completion of incubation period, growth of salt and temperature tolerant rhizobacterial isolates were observed and rated as +++: full growth, ++ : scanty growth +: growth and - : no growth. The full growth rhizobacterial isolates were selected for further studies.

Assessment of salt and temperature tolerant rhizobacteria on different temperature range.

To demonstrate the temperature tolerance also examined their ability to tolerate heat stress. In brief, pouring of 1 ml 24 hour old bacterial culture in 10 ml of six test tube of Ashby’s Mannitol broth medium out of which one kept as a control and remaining five tested for temperature tolerance by placing the test tube in water bath for 10 minute at different temperature 45, 50, 55, 60 and 65⁰C consequently place that test tube incubated in incubator at 28 ± 2 ⁰c for 48 hour. As a result of that culture colour change observed at selective temperature, after that take one ml of each temperature test tube broth and grow on Ashby’s Mannitol agar media and observed growth by spreading each strain on Ashby’s Mannitol agar medium and then plates were incubated in incubator at 28 ± 2 ⁰c for 48 hour.

Table 1

Assessment of salt and temperature tolerance rhizobacteria from rhizosphere soil of tomato on different NaCl concentrations
Isolate No.	Salt tolerance limit (NaCl %)										Class
Isolate No.	0.15	0.3	0.6	0.9	3.0	6.0	9.6	10.8	16.0	25.0
STT-A1	+++	+++	++	++	++	--	--	--	--	--	Tolerant
STT-A2	++	+	--		--	--	--	--	--	--	Moderately sensitive
STT-A3	+++	+++	++	+	--	--	--	--	--	--	Moderately tolerant
STT-A4	++	+	--	--	--	--	--	--	--	--	Moderately sensitive
STT-A5	++	+	--	--	--	--	--	--	--	--	Moderately sensitive
STT-A6	++	+	--	--	--	--	--	--	--	--	Moderately sensitive
STT-A7	+++	+++	++	+	--	--	--	--	--	--	Moderately tolerant
STT-A8	+++	+++	+++	+++	+++	++	--	--	--	--	Extremely tolerant
STT-A9	+++	+++	++	+	--	--	--	--	--	--	Moderately tolerant
STT-A10	+++	+++	+++	+++	+++	++	--	--	--	--	Extremely tolerant
STT-A11	++	+	--	--	--	--	--	--	--	--	Moderately sensitive
STT-A12	+++	+++	+++	+++	+++	++	--	--	--	--	Extremely tolerant
STT-A13	++	+	--	--	--	--	--	--	--	--	Moderately sensitive
STT-A14	++	+	--	--	--	--	--	--	--	--	Moderately sensitive
STT-A15	++	+	--	--	--	--	--	--	--	--	Moderately sensitive
STT-A16	+++	+++	++	--	--	--	--	--	--	--	Moderately sensitive
STT-A17	+++	+++	+++	+++	+++	++	--	--	--	--	Extremely tolerant
STT-A18	+++	+++	++	+	--	--	--	--	--	--	Moderately tolerant
STT-A19	++	+	--		--	--	--	--	--	--	Susceptible
STT-A20	+++	+++	++	+	--	--	--	--	--	--	Moderately tolerant
STT-A21	++	+	--	--	--	--	--	--	--	--	Moderately sensitive e
STT-A22	++	+	--	--	--	--	--	--	--	--	Moderately sensitive
STT-A23	++	+	--	--	--	--	--	--	--	--	Moderately sensitive
STT-A24	+++	+++	+++	+++	+++	++	--	--	--	--	Extremely
											tolerant
STT-A25	++	+	--	--	--	--	--	--	--	--	Moderately sensitive
STT-A26	++	+	--	--	--	--	--	--	--	--	Moderately sensitive
STT-A27	+++	+++	+++	+++	+++	++	--	--	--	--	Extremely tolerant
STT-A28	+++	+++	+++	+++	+++	++	--	--	--	--	Extremely tolerant
STT-A29	++	+	--	--	--	--	--	--	--	--	Moderately sensitive
STT-A30	+++	+++	++	+	--	--	--	--	--	--	Moderately tolerant
STT-A31	+++	+++	++	--	+	--	--	--	--	--	Tolerant
STT-A32	++	+	--		--	--	--	--	--	--	Moderately sensitive
STT-A33	+++	+++	++	+	--	--	--	--	--	--	Moderately tolerant
STT-A34	+++	+++	++	+	--	--	--	--	--	--	Moderately tolerant
STT-A35	++	+	--	--	--	--	--	--	--	--	Moderately sensitive
STT-A36	+++	+++	++	++	++	--	--	--	--	--	Tolerant
STT-A37	++	+	--	--	--	--	--	--	--	--	Moderately sensitive
STT-A38	++	+	--	--	--	--	--	--	--	--	Moderately sensitive
STT-A39	+++	+++	+++	+++	+++	++	--	--	--	--	Extremely tolerant
STT-A40	++	+	--	--	--	--	--	--	--	--	Moderately sensitive
STT-A41	+++	+++	++	+	--	--	--	--	--	--	Moderately tolerant
STT-A42	++	+	--	--	--	--	--	--	--	--	Moderately sensitive
STT-A43	++	+	--	--	--	--	--	--	--	--	Moderately sensitive
STT-A44	++	+	--	--		--	--	--	--	--	Moderately sensitive
STT-A45	++	+	--	--		--	--	--	--	--	Moderately sensitive
STT-A46	+++	+++	+++	+++	+++	++	--	--	--	--	Extremely tolerant

++ +: thick colony, + +: flattened, +: growth, -: No growth

extremely sensitive :tolerance limit less than 0.15% salt

Moderately sensitive :tolerance limit 0.15–0.60% salt

Moderately tolerant :tolerance limit 0.90–1.50% salt

Tolerant:tolerance limit 2.10–3.60% salt

Extremely tolerant :tolerance limit more than 5.40% salt (Cardoso et al., 2014).

Table 2

Classification of salt and temperature tolerance rhizobacteria based on salt tolerance limit
Salt tolerance limit	Isolate No.	No. of isolates	Location of Sample collection
less than 0.15% salt (Extremely sensitive)	_	0	_
0.15–0.60% salt (Moderat ely sensitive)	STT-A2,STT-A3,STT-A4,STT- A5,STT-A6,STT-A11,STT- A13,STT-A14,STT-A15,STT- A16,STT-A19,STT-A21,STT- A22,STT-A23,STT-A25,STT- A26,STT-A29,STT-A32,STT- A35,STT-A37,STT-A38,STT- A40,STT-A42,STT-A43,STT- A44,STT-A45	26	Khandgao village and Chorpada khandgao village Sangamner Ahmednagar, Satara and Pune,
0.9–1.5% salt (Moderately tolerant)	STT-A1,STT-A7,STT-A9,STT-A18,STT- A20,STT-A30,STT-A33,STT-A34,STT- A41	9	Khandgao village and Chorpada khandgao village, Sangamner, Ahmednagar, Anjaneri Nashik and Satara
2.1–3.6% salt (Tolerant)	STT-A31,STT-A36	02	Anjaneri nashik
more than 5.4% salt (Extremely tolerant)	STT-A8,STT-A10,STT-A12,STT- A17,STT-A24,STT-A27,STT- A28,STT-A39,STT-A46	9	Khandgao village and Chorpada khandgao village, Sangamner tehsil Ahmednagar, Anjaneri Nashik and Satara

Total forty six (46) salt and temperature tolerance rhizobacterial pure culture isolates were obtained. These rhizobacterial isolates were further assessed for tolerance ability on different NaCl cocentration, MgCl₂ concentrations and pH level by adjusting NaCl cocentration, MgCl₂ concentrations and pH level of culture media.

Among forty six isolates, nine isolates were extremely salt tolerant (more than 5.4% salt tolerance) limit. Two isolates were tolerant to NaCl concentration (2.10–3.6%). Nine isolates were moderately tolerant to NaCl concentration (0.09–1.50%) and twenty six isolates were moderately sensitive to NaCl concentration (0.15–0.60%) (Table 1, Plate 1, 2 and 3). Classification of salt and temperature tolerance of rhizobacteria was done on the basis of salt tolerance limit (Table 2). Similar results were found in investigations of Kumar et al., (2014) certain bacterial strains exhibited tolerance to high salinity (up to 7% NaCl). Furthermore stress tolerance was comparatively lower in Pseudomonas species as opposed to Bacillus strains, largely due to the formation of endospores by the Bacillus isolates. In a related study by Patel et al., (2017), they found that out of the isolates tested, 29 (43.0%) were able to thrive in the presence of 15 g of NaCl per 100 ml (w/v).The results are in agreement with observations of Damodarachari et al., (2018) conducted a study in which they found that five isolates (BS 1, BS 3, BS 14, BS 18, BS 42) demonstrated resilience to NaCl concentrations ranging from 1.5–20%. Similarly, Sharma et al., (2021) also yielded comparable results. In their study, isolates HB6P2 and HB6J2 demonstrated the highest level of salt tolerance, with a peak tolerance at 10% salt concentration. This was followed by isolates HB4A1, HB4N3, and HB8P1, which also exhibited notable tolerance to salts. Kumari et al., (2022) presented similar findings in their research. They identified isolates E-2, T-2, and T-1, which displayed growth at salt (NaCl) concentrations of 7%, 6%, and 6%, respectively. Similar results were found in investigations of several scientists Date 1982; Rai and Prasad 1986; Paknikar 1987; Zahran et al.1995,1999; Patil et al. 2014; Kumari and Khanna 2015 and Khan et al., 2019.

Table 3

Assessment of rhizobacteria for salt and temperature tolerance on different MgCl₂ concentration on medium
Isolate No.	MgCl₂ concentration (%)
Isolate No.	0.5	1.0	2.0	3.0	4.0
STT-A1	++	++	++	++	--
STT-A2	++	++	--	--	--
STT-A3	++	++	--	--	--
STT-A4	++	++	--	--	--
STT-A5	++	++	--	--	--
STT-A6	++	++	--	--	--
STT-A7	++	++	--	--	--
STT-A8	++	++	++	++	++
STT-A9	++	++	--	--	--
STT-A10	++	++	++	++	++
STT-A11	++	++	--	--	--
STT-A12	++	++	++	++	++
STT-A13	++	++	--	--	--
STT-A14	++	++	--	--	--
STT-A15	++	++	--	--	--
STT-A16	++	++	--	--	--
STT-A17	++	++	++	++	++
STT-A18	++	++	--	--	--
STT-A19	++	++	--	--	--
STT-A20	++	++	--	--	--
STT-A21	++	++	++	--	--
STT-A22	++	++	++	--	--
STT-A23	++	++	++	--	--
STT-A24	++	++	++	++	++
STT-A25	++	++	++	--	--
STT-A26	++	++	++	--	--
STT-A27	++	++	++	++	++
STT-A28	++	++	++	++	++
STT-A29	++	++	++	--	--
STT-A30	++	++	++	--	--
STT-A31	++	++	++	--	--
STT-A32	++	++	++	--	--
STT-A33	++	++	++	--	--
STT-A34	++	++	++	--	--
STT-A35	++	++	++	--	--
STT-A36	++	++	++	--	--
STT-A37	++	--	--	--	--
STT-A38	++	--	--	--	--
STT-A39	++	++	++	++	++
STT-A40	++	--	--	--	--
STT-A41	++	--	--	--	--
STT-A42	++	--	--	--	--
STT-A43	++	--	--	--	--
STT-A44	++	--	--	--	--
STT-A45	++	--	--	--	--
STT-A46	++	++	++	++	--

Among forty six isolates, eight salt and temperature tolerance of rhizobacteria were showed growth (Table 3, Plate 4, 5 and 6) upto 4.0% MgCl₂ concentration. Seven isolates showed growth upto 3.0% MgCl₂ concentration. Four salt and temperature tolerance rhizobacteria showed growth upto 2.0% MgCl₂ concentration. Seven isolates showed growth upto 1.0% MgCl₂ concentration and twenty salt and temperature tolerance rhizobacteria showed growth upto 0.5% MgCl₂ concentration. The results are in agreement with observations of Pagare et al., (2018) out of thirty three isolates, twenty isolates showed growth at MgCl₂ concentration of 0.75%, four isolates showed growth at 0.50% MgCl₂ concentration and nine isolates showed growth at 0.40% MgCl₂ concentration.

Table 4

Assessment of salt and temperature tolerant rhizobacteria at different pH level.
Isolate No.	PH
Isolate No.	7.0	8.0	8.5	9.0
STT-A1	++	--	--	--
STT-A2	++	--	--	--
STT-A3	++	--	--	--
STT-A4	++	--	--	--
STT-A5	++	--	--	--
STT-A6	++	--	--	--
STT-A7	++	--	--	--
STT-A8	++	++	++	++
STT-A9	++	--	--	--
STT-A10	++	++	++	++
STT-A11	++	--	--	--
STT-A12	++	++	++	++
STT-A13	++	--	--	--
STT-A14	++	--	--	--
STT-A15	++	--	--	--
STT-A16	++	--	--	--
STT-A17	++	++	++	++
STT-A18	++	--	--	--
STT-A19	++	--	--	--
STT-A20	++	--	--	--
STT-A21	++	--	--	--
STT-A22	++	--	--	--
STT-A23	++	--	--	--
STT-A24	++	++	++	++
STT-A25	++	--	--	--
STT-A26	++	--	--	--
STT-A27	++	++	++	++
STT-A28	++	++	++	++
STT-A29	++	--	--	--
STT-A30	++	--	--	--
STT-A31	++	--	--	--
STT-A32	++	--	--	--
STT-A33	++	--	--	--
STT-A34	++	--	--	--
STT-A35	++	--	--	--
STT-A36	++	--	--	--
STT-A37	++	--	--	--
STT-A38	++	--	--	--
STT-A39	++	++	++	++
STT-A40	++	--	--	--
STT-A41	++	--	--	--
STT-A42	++	--	--	--
STT-A43	++	--	--	--
STT-A44	++	--	--	--
STT-A45	++	--	--	--
STT-A46	++	--	--	--

Out of forty six salt and temperature tolerance rhizobacterial isolates (Table 4, Plate 7, 8, 9 and 10) eleven isolates showed growth at pH 9.0, seventeen isolates showed growth at pH 8.50, ten isolates showed growth at pH 8.00 and eight isolates showed growth at pH 7.0.The results are in agreement with observations of Damodarachari et al., (2018) conducted a study in which they found that out of fourty four, four isolates (BS 1, BS 3, BS 14, BS 18) were capable of growing across a pH range of 4 to 12. The growth of rhizobacteria on different pH has been reported by many scientists (Damodarachari et al., 2018; Tsegaye et al., 2019; Jianyang et al., 2020).

Table 5

Assessment of salt and temperature tolerance rhizobacteria at different temperature ranges.
Isolates	Temperature
Isolates	Control	50⁰c	55⁰c	60⁰c	65⁰c	Colour developing temperature
STT-A8	+++	+++	+++	+++	---	Greenish-65⁰c
STT-A10	+++	+++	+++	+++	---	Colourless 65⁰c
STT-A12	+++	+++	+++	+++	---	Colourless 65⁰c
STT-A24	+++	+++	---	---	---	Light Yellow 55⁰c
STT-A27	+++	+++	---	---	---	Pale yellow 55⁰c
STT-A39	+++	+++	---	---	---	Pale yellow 55⁰c

Out of the forty six isolates, six isolates selected for further studies related to temperature tolerance( Table 5, Plate 11 and 12), out of six salt and temperature tolerant isolates, three isolates (STT-A8, STT-A10 and STT-A12) showed temperature tolerance to 60⁰c and three isolates (STT-A24, STT-A27 and STT-A39) showed temperature tolerance to 50⁰c. Similar results were found in investigations of Kumar et al., (2014) conducted research indicating that certain bacterial strains exhibited tolerance to high temperatures (up to 50°C). In a related study by Patel et al., (2017) 40 isolates (58.20%) displayed the capacity to tolerate high temperatures, withstanding up to 70°C.The results are in agreement with observations of Damodarachari et al., (2018) conducted a study in which they found that six isolates (BS 10, BS 14, BS 18, BS 27, BS 37, BS 43) exhibited tolerance to temperatures spanning from 20°C to 50°C.

In the present study isolation, selection and characterization of salt and temperature tolerant rhizobacteria have been studied. Based on the results obtained from this study all the four isolates STT-A8, STT-A12, STT-A24, STT-A39 can tolerate NaCl 6%, MgCl₂ 4% and pH 9.0. Pseudomonas furukawaii STT-A8, Achromobacter sp. STT-A12 Shown temperature tolerance limit at 60⁰c and Agrobacterium pusense STT-A24 and Agrobacterium pusense STT-A39 shown temperature tolerance limit at 50⁰c. In evaluating the results obtained from this research study, it can be deduced that all the four plant growth promoting salt and temperature tolerant rhizobacteria able to survive in saline soil and can provide fertilizer to plants.

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Plates 1 to 12 are available in the Supplementary Files section.

No competing interests reported.

platessatyam2research01.10.23.docx

Isolation and Characterization of Salt and Temperature Tolerant Plant Growth Promoting Rhizobacteria for Enhancing Tomato Plant Growth in Saline Environments

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Abstract

Introduction

Material and methods

Collection of rhizosphere soil samples of tomato

Serial dilution

Result and Discussion

Conclusion

References

Plates

Additional Declarations

Supplementary Files

Status:

Version 1